Figure 1

Distribution of L-whirlin in the developing and mature hair bundles of the mouse cochlea. (A) Schematic representation of the L-whirlin and S-whirlin isoforms. Red lines indicate the positions of the segments used to produce antibodies against L-whirlin and against both whirlin isoforms. (B) Whirlin immunolocalization in developing hair bundles. Left and middle panels: confocal images of OHC and IHC hair bundles stained for either L-whirlin (anti-L-whirlin NTR antibody; left panel, green) or both whirlin isoforms (anti-whirlin-PDZ3 antibody; middle panel, green) and for actin (red) on P6. The immunoreactivity is detected at the same places with the two antibodies, namely at the tips of OHC and IHC tall stereocilia (arrowheads), and in the ankle-link region (AL) of all stereocilia (white arrows). Right panel: confocal images of OHCs and IHCs immunolabeled for the ankle-link protein ADGRV1 (green; white arrows) on P6. Scale bar: 2 µm. (C) Whirlin immunolocalization in mature hair bundles. Left panel: Scanning electron micrograph of OHC and IHC hair bundles immunostained for L-whirlin on P16. Protein A conjugated to 15 nm-diameter gold particles was used for detection. The labeling is detected only at the tips of OHC tall stereocilia (arrowheads). Middle and right panels: confocal microscopy immunolabeling of L-whirlin (middle panel) and of both whirlin isoforms (right panel) on P20. The labeling is present at the tips of OHC and IHC tall stereocilia (arrowheads) with the two antibodies. Scale bar: 2 µm.